Heart disease is the leading cause of death in humans, and myocarditis is a predominant cause of heart failure in young adolescents. Patients affected with myocarditis can develop dilated cardiomyopathy (DCM), a common reason for heart transplantation, which to date is the only viable option for combating DCM. Myocarditis/DCM patients show antibodies to coxsackievirus B3 (CVB) and cardiac antigens, suggesting a role for CVB-mediated autoimmunity in the disease pathogenesis, but a direct causal link remains to be determined clinically. The long-term goal is to determine the autoimmune mechanisms of pathogens that commonly infect the cardiovascular system and initiate heart autoimmunity. The objective of this application is to determine whether chronic myocarditis that occurs in CVB infection due to an autoimmune response to cardiac antigens. The central hypothesis is that cardiac-specific CD4 T cells generated by mechanisms that involve molecular mimicry and epitope spreading mediate postinfectious myocarditis induced by CVB. This hypothesis will be tested in A/J mice that are highly susceptible to both autoimmune and viral myocarditis, the histologic features of which resemble those in human disease. The project's two specific aims are to: 1) Delineate the role of cross-reactive CD4 T cells in the mediation of CVB myocarditis. Panel of mimicry epitopes identified from the CVB proteome are capable of inducing pathogenic cross-reactive T cells for three cardiac antigens as evaluated by T cell proliferation assay and cytokine responses. [2) Investigate epitope spreading as an autoimmune mechanism of postinfectious myocarditis induced by CVB. Using major histocompatibility complex (MHC) class II dextramers for cardiac myosin heavy chain (Myhc)- 334-352, the team has demonstrated that myocarditis-susceptible A/J mice, show the generation of Myhc--specific T cells that transfer disease to naive mice. These observations provide a compelling rationale to test the hypothesis that CVB infection induces the generation of cardiac-reactive T cells with multiple antigen specificities through epitope spreading. The specific aims will be achieved by measurement of T cell responses, MHC class II dextramer staining, cytokine analysis, magnetic resonance microscopy imaging, active immunization and adoptive transfer experiments, and histology. The proposed approach is innovative, as it addresses the fundamental question of whether autoimmune response contributes to postinfectious myocarditis induced by CVB at the level of antigen specificity. The proposed research is significant, as it may provide a basis for future investigations into the role of cardic-reactive T cells that might be generated in cardiomyopathy patients as a result of exposure to CVB. These studies may then create opportunities to derive immunotherapies in the form of altered peptide ligands and tolerogens as antigen-specific treatment modalities, including personalized therapies, for DCM patients.]